The soluble methane menooxygenase system isolated from M. capsulatus (Bath) consists of a reductase (MMOR), coupling protein (MMOB), and a hydroxylase (MMOH). The 16.5 kDa MMOB component has a profound influence on the type of chemistry catalyzed at the active site of MMOH. In a still unknown mechanism, MMOB directs the evolution of a series of oxygen intermediates that lead to the insertion of an atom of molecular oxygen into a CH bond of methane. In the absence of MMOB, MMOH still accepts electrons from substrate specificity of MMOH. This broad substrate specificity makes MMOH valuable system in both aqueous phase organic synthesis, and in the detoxification of contaminated environmental sites. High resolution homonuclear and heteronuclear NMR experiments will be used to solve the solution structure of MMOB. The three dimensional NMR structure will provide greater insight into the way in which MMOB tunes the reactivity potentials of the MMOH iron center. Searches based on the complementarity of the topologies and electrostatic surface potentials of MMOH and MMOB will be used in conjunction with chemical cross-linking data to identify probable MMOB binding sites on the surface of MMOH. Proposed binding sites will be confirmed through titration calorimetric studies involving synthetic peptides representative of secondary structural motifs in interface region. Individual amino acid side chains involved in the complex formation will be identified in high resolution NMR studies of complexes of MMOB with synthetic peptides. Amino acid side chains found to be involved in the binding interaction will be sequentially mutated to alanine. The relative contribution of individual amino acid side chains the macroscopic stability of the complexes formed between MMOB and MMOH will be measured based on calorimetric measurements.